The present invention generally relates to an improved method and apparatus for the analysis of gas phase ions by field asymmetric ion mobility spectrometry and by mass spectrometry. The apparatus and methods for sample handling and analysis described herein are enhancements of the techniques referred to in the literature relating to mass spectrometry—an important tool in the analysis of a wide range of chemical compounds. Specifically, mass spectrometers can be used to determine the molecular weight of sample compounds. The analysis of samples by mass spectrometry consists of three main steps—formation of gas phase ions from sample material, mass analysis of the ions to separate the ions from one another according to ion mass, and detection of the ions. A variety of means and methods exist in the field of mass spectrometry to perform each of these three functions. The particular combination of the means and methods used in a given mass spectrometer determine the characteristics of that instrument.
To mass analyze ions, for example, one might use magnetic (B) or electrostatic (E) analysis, wherein ions passing through a magnetic or electrostatic field will follow a curved path. In a magnetic field, the curvature of the path will be indicative of the momentum-to-charge ratio of the ion. In an electrostatic field, the curvature of the path will be indicative of the kinetic energy-to-charge ratio of the ion. If magnetic and electrostatic analyzers are used consecutively, then both the momentum-to-charge and kinetic energy-to-charge ratios of the ions will be known and the mass of the ion will thereby be determined. Other well known mass analyzers are the quadrupole (Q), the ion cyclotron resonance (ICR), the time-of-flight (TOF), and the Paul ion trap analyzers. More recently, linear quadrupole ion traps [J. Schwartz, M. Senko, and J. Syka, J. Am. Soc. Mass Spectrom. 13, 659(2002); J. Hager, Rapid Commun. Mass Spectrom. 16, 512(2002)] have become more wide spread. And a new analyzer, the orbitrap, based on the Kingdon trap [K. Kingdon, Phys. Rev. 21, 408(1923)] was recently described by A. Makarov [Q. Hu et al., J Mass Spectrom. 40, 430(2005)]. Any form of mass analyzer may be used in conjunction with the means and method described here.
Before mass analysis can begin, gas phase ions must be formed from a sample material. If the sample material is sufficiently volatile, ions may be formed by electron ionization (EI) or chemical ionization (CI) of the gas phase sample molecules. Alternatively, for solid samples (e.g., semiconductors, or biological materials), ions can be formed by desorption and ionization of sample molecules by bombardment with high energy particles. Further, Secondary Ion Mass Spectrometry (SIMS), for example, uses keV ions to desorb and ionize sample material. In the SIMS process a large amount of energy is deposited in the analyte molecules, resulting in the fragmentation of fragile molecules. This fragmentation is undesirable in that information regarding the original composition of the sample (e.g., the molecular weight of sample molecules) will be lost.
For more labile, fragile molecules, other ionization methods now exist. The plasma desorption (PD) technique was introduced by Macfarlane et al. (R. D. Macfarlane, R. P. Skowronski, D. F. Torgerson, Biochem. Biophys. Res Commoun. 60 (1974) 616)(“McFarlane”). Macfarlane discovered that the impact of high energy (MeV) ions on a surface, like SIMS would cause desorption and ionization of small analyte molecules. However, unlike SIMS, the PD process also results in the desorption of larger, more labile species (e.g., insulin and other protein molecules).
Additionally, lasers have been used in a similar manner to induce desorption of biological or other labile molecules. See, for example, Cotter et al. (R. B. VanBreeman, M. Snow, R. J. Cotter, Int. J. Mass Spectrom. Ion Phys. 49 (1983) 35; Tabet, J. C.; Cotter, R. J., Tabet, J. C., Anal. Chem. 56 (1984) 1662; or R. J. Cotter, P. Demirev, I. Lys, J. K. Olthoff, J. K.; Lys, I.: Demirev, P.: Cotter et al., R. J., Anal. Instrument. 16 (1987) 93). Cotter modified a CVC 2000 time-of-flight mass spectrometer for infrared laser desorption of non-volatile biomolecules, using a Tachisto (Needham, Mass.) model 215G pulsed carbon dioxide laser. The plasma or laser desorption and ionization of labile molecules relies on the deposition of little or no energy in the analyte molecules of interest.
The use of lasers to desorb and ionize labile molecules intact was enhanced by the introduction of matrix assisted laser desorption ionization (MALDI) (K. Tanaka, H. Waki, Y. Ido, S. Akita, Y. Yoshida, T. Yoshica, Rapid Commun. Mass Spectrom. 2 (1988) 151 and M. Karas, F. Hillenkamp, Anal. Chem. 60 (1988) 2299). In the MALDI process, an analyte is dissolved in a solid, organic matrix. Laser light of a wavelength that is absorbed by the solid matrix but not by the analyte is used to excite the sample. Thus, the matrix is excited directly by the laser, and the excited matrix sublimes into the gas phase carrying with it the analyte molecules. The analyte molecules are then ionized by proton, electron, or cation transfer from the matrix molecules to the analyte molecules. This process (i.e., MALDI) is typically used in conjunction with time-of-flight mass spectrometry (TOFMS) and can be used to measure the molecular weights of proteins in excess of 100,000 Daltons.
Atmospheric Pressure Ionization (API) includes a number of ion production means and methods. Among these are atmospheric pressure chemical ionization (APCI), atmospheric pressure photoionization (APPI), electrospray ionization (ESI), and desorption electrospray ionization (ESI). Typically, analyte ions are produced from liquid solution at atmospheric pressure. ESI, one of the more widely used methods, was first suggested for use with mass spectrometry by Dole et al. (M. Dole, L. L. Mack, R. L. Hines, R. C. Mobley, L. D. Ferguson, M. B. Alice, J. Chem. Phys. 49, 2240, 1968). In the electrospray technique, analyte is dissolved in a liquid solution and sprayed from a needle. The spray is induced by the application of a potential difference between the needle and a counter electrode. The spray results in the formation of fine, charged droplets of solution containing analyte molecules. In the gas phase, the solvent evaporates leaving behind charged, gas phase, analyte ions. This method allows for very large ions to be formed. Ions as large as 1 MDa have been detected by ESI in conjunction with mass spectrometry (ESMS).
In addition to ESI, many other ion production methods might be used at atmospheric or elevated pressure. For example, MALDI has recently been adapted by Laiko et al. to work at atmospheric pressure (Victor Laiko and Alma Burlingame, “Atmospheric Pressure Matrix Assisted Laser Desorption”, U.S. Pat. No. 5,965,884, and Atmospheric Pressure Matrix Assisted Laser Desorption Ionization, poster #1121, 4th International Symposium on Mass Spectrometry in the Health and Life Sciences, San Francisco, Aug. 25-29, 1998) and by Standing et al. at elevated pressures (Time of Flight Mass Spectrometry of Biomolecules with Orthogonal Injection+Collisional Cooling, poster #1272, 4th International Symposium on Mass Spectrometry in the Health and Life Sciences, San Francisco, Aug. 25-29, 1998; and Orthogonal Injection TOFMS Anal. Chem. 71(13), 452A (1999)). The benefit of adapting ion sources in this manner is that the ion optics (i.e., the electrode structure and operation) in the mass analyzer and mass spectral results obtained are largely independent of the ion production method used.
The elevated pressure MALDI source disclosed by Standing differs from what is disclosed by Laiko et al. Specifically, Laiko et al. disclose a source intended to operate at substantially atmospheric pressure. In contrast, the source disclosed by Standing et al. is intended to operate at a pressure of about 70 mtorr.
More recently, Takats et al. [Z. Takats, J. M. Wiseman, B. Gologan, and R. G. Cooks, Science 306, 471(2004)] introduced yet another atmospheric pressure ionization method known as desorption electrospray ionization (DESI). According to Takats et al., DESI is a method for producing ions from analyte on a surface. Electrosprayed charged droplets and ion of solvent are directed at the surface under study. The impact of the charged droplets on the surface results in the desorption and ionization of the analyte to form gas phase analyte ions.
Gas phase ions may be analyzed via any of the above described mass analyzers, via an ion mobility analyzer, or by a combination of mass and mobility analyzers. Ion mobility spectrometry (IMS) is a method whereby the “mobility” of analyte ions through a gas is measured under the influence of a static electric field. IMS is described in detail in the literature [see, for example, G. Eiceman and Z. Karpas, Ion Mobility Spectrometry (CRC. Boca Raton, Fla. 1994); and Plasma Chromatography, edited by T. W. Carr (Plenum, New York, 1984)]. At low electric field strengths—e.g. a few kilovolts per meter—the speed of analyte ions through a gas is measured. To start the measurement, ions are pulsed into the entrance of the mobility analyzer. Ions of a given mobility travel the length of the drift tube of the mobility analyzer at fixed velocity resulting from the balance in force between the electric field pushing ions forward and drag on the ions due to collisions with the gas. At the far end of the analyzer, the ions strike a detector and are detected. By measuring the time between the introduction of ions into the analyzer and the detection of the ions, the speed of the ions, and therefore their mobility can be determined.
At low field strengths, the mobility of an ion is a constant relating the speed of the ion to the strength of the electric field. However, at high electric field strengths, the mobility of the ions varies with electric field strength. This gives rise to field asymmetric ion mobility spectrometry (FAIMS)—an extension of IMS which takes advantage of the change in ion mobility at high field strengths.
FAIMS is based on an observation of Mason and McDaniel [W. McDaniel and Edward A. Mason, The mobility and diffusion of ions in gases, John Wiley & Sons, 1973] who found that the mobility of an ion varies with the applied electric field strength. Above an electric field to gas density ratio (E/N) of 40 Td (E>10,700 V/cm at atmospheric pressure) the mobility coefficient K(E) has a non-linear dependence on the field. This dependence is believed to be specific for each ion species. A coefficient “a” describes the change in mobility as a function of field strength and is defined as the fractional change in mobility when comparing a high field strength condition to a low field strength condition. An α value of 0.1, for example, represents an increase of 10% in the ion's mobility whereas an α value of −0.1 represents a decrease of 10% in the ion's mobility.
FAIMS is described in detail in the literature [I. Buryakov, E. Krylov, E. Nazarov, and U. Rasulev, Int. J. Mass Spectrom. Ion Phys. 128. 143 (1993); D. Riegner, C. Harden, B. Carnahan, and S. Day, Proceedings of the 45th ASMS Conference on Mass Spectrometry and Allied Topics, Palm Springs, Calif., Jun. 1-4, 1997, p. 473; B. Carnahan, S. Day, V. Kouznetsov, M. Matyjaszczyk, and A. Tarassov, Proceedings of the 41st ISA Analysis Division Symposium, Framingham, Mass., Apr. 21-24, 1996, p. 85; and B. Carnahan and A. Tarassov, U.S. Pat. No. 5,420,424]. FAIMS devices measure the difference in the mobility of an ion at high field relative to its mobility at low field. That is, the ions are separated on the basis of the compound dependent behavior of mobility as a function of electric field strength. In the prior art FAIMS devices such as described in U.S. Pat. No. 6,972,407, herein incorporated by reference, two parallel, planar conducting electrodes are used to generate an electric field in which analyte ions are to be analyzed. The analyte ions are entrained in a carrier gas which moves at high velocity (several meters per second) perpendicular to the electric field—i.e. parallel to the surface of the planar conducting electrodes. Applying the appropriate potentials to the “top” and “bottom” electrodes will result in the filtering of ions on the basis of their α value. As described in the prior art literature, a rectangular waveform having repeatedly a high potential and then a low potential is applied between the electrodes. For a relatively short period of time, a high potential is applied between the electrodes and then for a longer period of time a relatively low potential of opposite polarity is applied. The magnitude of the potentials and the duration of their application are such that the time averaged potential difference is zero. During the application of the high potential ions will drift laterally with a mobility K(1+α). During the application of the low potential of opposing polarity, the ions will drift laterally with a mobility K and in the opposite direction to that when the high potential was applied. Applying an additional DC “compensation voltage” between the electrodes allows the selection of ions of a given mobility difference to be transmitted.
In recent years, IMS and FAIMS spectrometers have been combined with mass spectrometry. In U.S. Pat. No. 5,905,258 Clemmer and Reilly combine IMS with a time of flight mass spectrometer (TOFMS). This provided for a first analysis of the ions by IMS followed by a second analysis via TOFMS. Ultimately, this yields a two dimensional plot containing both the mobility and mass of the ions under investigation. The advantages of this type of combined analyzer over a mass spectrometer alone are described in detail in the literature [C. S. Srebalus et al., Anal. Chem. 71(18), 3918(1999); J. A. Taraszka et al., J. Proteom. Res. 4, 1223(2005); R. L. Wong, E. R. Williams, A. E. Counterman, and D. Clemmer, J. Am. Soc. Mass Spectrom. 16, 1009(2005)] and include the separation of chemical background from analyte species for an improved signal-to-noise ratio (S/N), and the separation of ions based on compound class or charge state for easier mass spectral interpretation.
Similarly, in U.S. Pat. No. 6,504,149, for example, Guevremont et al. combine a FAIMS device with a mass spectrometer. As detailed in the literature, a combined FAIMS mass spectrometer has similar advantages as an IMS mass spectrometer [A. Shvartsburg, K. Tang, R. Smith, J. Am. Soc. Mass Spectrom. 16, 2(2005); D. A. Barnett, B. Ells, R. Guevremont, and R. W. Purves, J. Am. Soc. Mass Spectrom. 13, 1282(2002)]. For example, a combined FAIMS mass spectrometer has an improved signal-to-noise ratio over a mass spectrometer alone because the FAIMS device can filter away the chemical background.
In the instrument disclosed by Guevremont, the FAIMS device is imposed between the ion production means and the inlet to the mass spectrometer. That is, a FAIMS device may be added to a preexisting mass spectrometer by moving the ion production means and placing the FAIMS apparatus between the inlet to the mass spectrometer and the ion production means. As a result, the transmission efficiency of analyte ions from the ion production means to the mass analyzer is reduced. Although the S/N in the mass spectra produced may be improved over a mass spectrometer alone, the imposition of the FAIMS device nonetheless leads to a reduced sensitivity.
Furthermore prior art FAIMS devices have been employed as peripheral or add-on devices when used in conjunction with mass spectrometers [B. Ells et al, Anal. Chem. 71, 4747(1999)]. That is, prior art FAIMS devices are not highly integrated with mass spectrometers and users or technicians must mount and demount the FAIMS device in order to run tandem FAIMS/MS or MS—only experiments, respectively. That is, if a prior art FAIMS apparatus is implemented on a mass spectrometer then it must be kept in operation in order to be of any benefit. In order to observe the entire range of ions being generated by the ion production means—i.e. without filtering via FAIMS—the asymmetric waveform must be turned off. However, the presence of the FAIMS device between the ion source and the mass spectrometer—even if deactivated—would represent a loss of ion transmission efficiency and therefore a loss in sensitivity. Thus, in a practical operation, the FAIMS apparatus must be implemented between the ion production means and the inlet of the mass spectrometer when FAIMS filtering is desired and the FAIMS apparatus must be removed again when FAIMS filtering is not desired. This added complexity discourages users from adopting FAIMS in combination with mass spectrometry.
If gas phase ions produced via an API method are to be analyzed in a mass spectrometer, they must first be transported from the ionization region through regions of differing pressures and ultimately to a mass analyzer for subsequent analysis (e.g., via time-of-flight mass spectrometry (TOFMS), Fourier transform mass spectrometry (FTMS), etc.). In some prior art sources, this was accomplished through use of a small orifice between the ionization region and the vacuum region. In conventional instruments, the orifice is circular, however, in U.S. Pat. No. 7,339,166, Tang et al. recognize that the ion transmission efficiency between FAIMS analyzers of planar geometry and mass analyzers at reduced pressures can be improved by changing the shape of the orifice or by using a multitude of orifices. Specifically, a “ . . . conductance limit aperture having the geometry of a rectangle . . . provides a more efficient coupling of planar . . . FAIMS to downstream stages . . . . ”
In other prior art, metal or dielectric capillaries have been used to transmit ions entrained in a carrier gas from a high pressure ion production region into the vacuum chamber of mass spectrometers—see, for example, Fenn et al., U.S. Pat. No. 4,542,293 and Whitehouse et al., U.S. Pat. No. 5,844,237. In U.S. Pat. No. 6,777,672, incorporated herein by reference, Park describes a multiple section capillary for interfacing various ion production means and for transporting ions into the vacuum chamber of a mass spectrometer.
An example of a prior art transfer capillary is shown in FIG. 1. As depicted, capillary 7 comprises a generally cylindrical glass tube 2 having an internal bore 4. The ends of capillary 7 include a metal coating (e.g., platinum, copper, etc.) to form conductors 5 which encompass the outer surface of capillary 7 at its ends, leaving a central aperture 6 such that the entrance and exit to internal bore 4 are left uncovered. Conductors 5 may be connected to electrical contacts (not shown) in order to maintain a desired space potential at each end of capillary 7. In operation, a first electrode (one of conductors 5) of capillary 7 may be maintained at an extreme negative potential (e.g. −4,500V). This first electrode 5 acts as the entrance end of the capillary and resides at near atmospheric pressure. Positively charged analyte ions formed in the atmospheric pressure ion production region are attracted to the first electrode 5 and are entrained in the gas flow into the capillary. A second electrode (the other of conductors 5), acts as the exit end of the capillary and resides at the pressure of the first vacuum region of the mass spectrometer. This second of conductors 5 may form the first stage of a multi-stage lensing system for the final direction of the ions to the spectrometer, and may be maintained at a positive potential (e.g., 160 volts).
Importantly, ions are carried through the transfer capillary by entrainment in gas which is pumped from the ion production region, through the capillary, into the first vacuum region of the mass spectrometer. Typically, the gas pressure at the capillary inlet is about one atmosphere whereas the pressure at the capillary outlet, into the first pumping region, is between one and three millibar. Under these conditions, the velocity of the gas in the capillary is about 100 m/s. It is the “force” associated with this high velocity gas that is able to drive the ions away from the electrically attractive potential at the capillary entrance and towards the electrically repulsive potential at the capillary exit.
In other prior art, Glish et al. (U.S. Pat. No. 6,703,611) and Prior et al. (U.S. Pat. No. 6,455,846) independently describe transfer capillaries having flared entrances. According to Glish, the value of their flared capillary is that it “has increased positional alignment tolerances and . . . is capable of both single and multiple nanoelectrospray and standard electrospray ionization.” According to Prior, their flared capillary “provides greater efficiency in the transmission of gaseous ions from an ion source situated in a region of relatively high pressure, to the interior of a device maintained at a relatively low pressure.”
Finally, in U.S. Pat. No. 7,598,488, incorporated herein by reference, Park describes the integration of a FAIMS analyzer into an ion transfer capillary. In one embodiment, the transfer capillary is a multisection capillary having the FAIMS analyzer integrated in a first section of the capillary and having a union by which the first capillary section can be removably joined with a second section of the capillary. According to Park, “the transmission efficiency of analyte ions [through a FAIMS analyzer integrated into the ion transfer capillary] is improved over prior art FAIMS devices”
However, the prior art does not describe any means or mechanism that provides a smooth transition between a FAIMS analyzer in a first section of capillary and a second section of the capillary. For example, a FAIMS analyzer in a first capillary section having a first geometric cross section—for example, rectangular—joined with a second section of capillary having a second cross section—for example, round—will result in a geometric discontinuity at the union between the two sections. Such a discontinuity leads, in prior art devices to turbulent gas flow, dead volumes, and a reduction in the transmission of ions from the outlet of the FAIMS analyzer into the downstream vacuum system and analyzers. Similarly, in a contiguous capillary, the cross section of the capillary in that part of the capillary which includes the FAIMS analyzer may differ from the remainder of the capillary. The geometric discontinuity between the FAIMS analyzer and the remainder of the capillary results in a loss of ion transmission.